Classifications

• • C—CHEMISTRY; METALLURGY
  • C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
  • C09B—ORGANIC DYES OR CLOSELY-RELATED COMPOUNDS FOR PRODUCING DYES, e.g. PIGMENTS; MORDANTS; LAKES
  • C09B3/00—Dyes with an anthracene nucleus condensed with one or more carbocyclic rings
  • C09B3/14—Perylene derivatives
• • C—CHEMISTRY; METALLURGY
  • C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
  • C09B—ORGANIC DYES OR CLOSELY-RELATED COMPOUNDS FOR PRODUCING DYES, e.g. PIGMENTS; MORDANTS; LAKES
  • C09B69/00—Dyes not provided for by a single group of this subclass
  • C09B69/10—Polymeric dyes; Reaction products of dyes with monomers or with macromolecular compounds
  • C09B69/101—Polymeric dyes; Reaction products of dyes with monomers or with macromolecular compounds containing an anthracene dye
• • C—CHEMISTRY; METALLURGY
  • C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
  • C09B—ORGANIC DYES OR CLOSELY-RELATED COMPOUNDS FOR PRODUCING DYES, e.g. PIGMENTS; MORDANTS; LAKES
  • C09B69/00—Dyes not provided for by a single group of this subclass
  • C09B69/10—Polymeric dyes; Reaction products of dyes with monomers or with macromolecular compounds
  • C09B69/101—Polymeric dyes; Reaction products of dyes with monomers or with macromolecular compounds containing an anthracene dye
  • C09B69/102—Polymeric dyes; Reaction products of dyes with monomers or with macromolecular compounds containing an anthracene dye containing a perylene dye
• • C—CHEMISTRY; METALLURGY
  • C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
  • C09B—ORGANIC DYES OR CLOSELY-RELATED COMPOUNDS FOR PRODUCING DYES, e.g. PIGMENTS; MORDANTS; LAKES
  • C09B69/00—Dyes not provided for by a single group of this subclass
  • C09B69/10—Polymeric dyes; Reaction products of dyes with monomers or with macromolecular compounds
  • C09B69/103—Polymeric dyes; Reaction products of dyes with monomers or with macromolecular compounds containing a diaryl- or triarylmethane dye
• • C—CHEMISTRY; METALLURGY
  • C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
  • C09B—ORGANIC DYES OR CLOSELY-RELATED COMPOUNDS FOR PRODUCING DYES, e.g. PIGMENTS; MORDANTS; LAKES
  • C09B69/00—Dyes not provided for by a single group of this subclass
  • C09B69/10—Polymeric dyes; Reaction products of dyes with monomers or with macromolecular compounds
  • C09B69/109—Polymeric dyes; Reaction products of dyes with monomers or with macromolecular compounds containing other specific dyes
• • G—PHYSICS
  • G01—MEASURING; TESTING
  • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
  • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
  • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
  • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
  • G01N33/58—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances
  • G01N33/582—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances with fluorescent label
• • G—PHYSICS
  • G01—MEASURING; TESTING
  • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
  • G01N2510/00—Detection of programmed cell death, i.e. apoptosis

Definitions

• Heteroalkylene refers to an alkylene group comprising at least one heteroatom (e.g., N, O or S).
• the heteroatom is within the alkylene chain (i.e., the heteroalkylene comprises at least one carbon-heteroatom- carbon bond.
• the heteroatom is at a terminus of the alkylene and thus serves to join the alkylene to the remainder of the molecule (e.g., M1-H-A-M2, where Ml and M2 are portions of the molecule, H is a heteroatom and A is an alkylene).
• Heteroalkenylene is a heteroalkylene comprising at least one carbon- carbon double bond.
• Heteroalkynylene is a heteroalkylene comprising at least one carbon-carbon triple bond. Unless stated otherwise specifically in the specification, heteroalkylene, heteroalkenylene and heteroalkynylene are optionally substituted.
• Heteroatomic in reference to a “heteroatomic linker” refers to a linker group consisting of one or more heteroatom.
• Nonlimiting examples of photostable visible dyes suitable for use in the compounds and methods of the invention include azo dyes, thioindigo dyes, quinacridone pigments, dioxazine, phthalocyanine, perinone, diketopyrrolopyrrole, quinophthalone, and truarycarbonium.
• perylene derivative is intended to include any substituted perylene that is visually detectable. However, the term is not intended to include perylene itself.
• anthracene derivative e.g., perylene, pyrene, anthracene or naphthalene derivative
• a derivative is an imide, bisimide or hydrazamimide derivative of perylene, anthracene, naphthalene, or pyrene.
• the visually detectable molecules of the invention are useful for a wide variety of analytical applications, such as biochemical and biomedical applications, in which there is a need to determine the presence, location, or quantity of a particular analyte (e.g., biomolecule).
• the invention provides a method for visually detecting a biomolecule, comprising: (a) providing a biological system with a visually detectable biomolecule comprising the compound of structure (I) linked to a biomolecule; and (b) detecting the biomolecule by its visible properties.
• R d is a cation
• M is a fluorescent or colored moiety. Any fluorescent and/or colored moiety known in the art and typically employed in colorimetric, UV, and/or fluorescent assays may be used. Examples of M moieties which are useful in various embodiments of the invention include, but are not limited to: Xanthene derivatives (e.g., fluorescein, rhodamine, Oregon green, eosin or Texas red); Cyanine derivatives (e.g., cyanine, indocarbocyanine, oxacarbocyanine, thiacarbocyanine or merocyanine); Squaraine derivatives and ring-substituted squaraines, including Seta, SeTau, and Square dyes; Naphthalene derivatives (e.g., dansyl and prodan derivatives); Coumarin derivatives; oxadiazole derivatives (e.g., pyridyloxazole, nitrobenzoxadiazole or benzoxadiazol
• M is cyclic.
• M is carbocyclic.
• M is heterocyclic.
• M at each occurrence, independently comprises an aryl moiety.
• the aryl moiety is multicyclic.
• the aryl moiety is a fused-multicyclic aryl moiety, for example which may comprise at least 3, at least 4, or even more than 4 aryl rings.
• M 1 is, at each occurrence, independently p-terphenyl, perylene, azobenzene, phenazine, phenanthroline, acridine, thioxanthrene, chrysene, rubrene, coronene, cyanine, perylene imide, or perylene amide or a derivative thereof.
• M is, at each occurrence, independently a coumarin dye, resorufin dye, dipyrrometheneboron difluoride dye, ruthenium bipyridyl dye, energy transfer dye, thiazole orange dye, polymethine or N-aryl-l,8-naphthalimide dye.
• said sample comprises cells, and some embodiments further comprise observing said cells by flow cytometry.
• the disclosure provides a method for visually detecting an analyte molecule, such as a biomolecule, comprising:
• the analyte molecule is a nucleic acid, amino acid or a polymer thereof (e.g., polynucleotide or polypeptide). In still more embodiments, the analyte molecule is an enzyme, receptor, receptor ligand, antibody, glycoprotein, aptamer or prion.
• Oligomer dyes were synthesized on an Applied Biosy stems 394 DNA/RNA synthesizer or on GE AKTA 10 OligoPilot on either 1 ⁇ or 10 ⁇ scales and possessed a 3 '-phosphate group.
• Dyes were synthesized directly on CPG beads or on polystyrene solid support. The dyes were synthesized in the 3 ' to 5' direction by standard solid phase DNA methods. Coupling methods employed standard ⁇ -cyanoethyl phosphoramidite chemistry conditions. Different number of "m" repeating units were incorporated by repeating the synthesis cycle the desired number of times with an appropriate phosphoramidite.
• Extinction coefficients ( ⁇ ) were determined by the total number of fluors (M moieties) in the dye construct, using 75,000 M “1 cm “1 for each fluorescein (F; read at 494nm); 34,500 for each pyrene (Y; read at 343nm); and 40,000 for each perylene (E; read at 440nm) present in the sequence. Spacers are presumed to have no effect on ⁇ .
• Dye compounds comprising a perylene moiety (“E") were prepared according to the above general procedures and their fluorescent emission properties determined at 50 nM and pH 9. The structure of these dyes is presented in Table 8.
• the general flow cytometry workflow includes the following steps:
• Mid-log phase Jurkat and Ramos or primary PBMC cells were induced into necrosis in vitro in order to form dead cells and cell debris.
• Cells were induced by heat stress by subjecting cultured cells first to 58°C for 3-5 minutes, then transferring them to ice for 10-15 minutes. Fresh cells were maintained by subjecting cultures to no treatment.
• Cells and dye e.g., bulk F(CioF) 9 or FINSA activated, amine reactive F(Ci 0 F) 9 SH, see Table 2
• Dyes were diluted serially in pure deionized water or sterile filtered phosphate buffered solution to a concentration range of 15,000-0.001 ⁇ in volumes of 25-200 ⁇ . Phosphate buffered saline was used for amine reactive dye (i.e. F(CioF) 9 SH).
• the dye solutions were then applied to cell preparations.
• the dye mixtures were then incubated for 30-40 minutes at 23-25°C. After incubation, cells were washed twice with buffer (containing protein, phosphate buffered saline, and metabolic inhibitors) by the addition of 600-800 ⁇ , ⁇ . Mixtures were then centrifuged (g x 350) for 5 minutes.

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• Organic Chemistry (AREA)
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• Life Sciences & Earth Sciences (AREA)
• Engineering & Computer Science (AREA)
• Molecular Biology (AREA)
• Biomedical Technology (AREA)
• Urology & Nephrology (AREA)
• Hematology (AREA)
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• Biotechnology (AREA)
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• Food Science & Technology (AREA)
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• General Physics & Mathematics (AREA)
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• Investigating, Analyzing Materials By Fluorescence Or Luminescence (AREA)
• Plural Heterocyclic Compounds (AREA)
• Investigating Or Analysing Materials By The Use Of Chemical Reactions (AREA)

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• 2016
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